Optimization And Antioxidant Activity Evaluation Of Egg White Peptides Prepared By Lactobacillus Fermentation-Assisted Enzymolysis

Eggs are inexpensive and rich in functional ingredients,which play an important role in people’s daily diets.With the increasing attention to health problems,natural bioactive proteins and bioactive peptides have received more and more attention.Among them,the egg white peptides in eggs have attracted widespread attention in the food and biopharmaceutical industries because of their antioxidant activity.At present,the production of egg white peptides is mostly prepared by enzymatic hydrolysis,and the antioxidant activity of the obtained egg white peptides is not good.How to efficiently and quickly prepare egg white peptides with good activity is one of the urgent problems that need to be solved in the food and biopharmaceutical industries.In this thesis,on the basis of the previous enzymatic hydrolysis,Lactobacillus acidophilus and Lactobacillus plantarum mixed bacteria were used to ferment the enzymatic products of egg white,and the response surface method was used to optimize the process conditions for fermentation-assisted preparation of egg white peptides.Then,the fermentation products prepared under optimal conditions were ultrafiltered,and different molecular weight ranges of egg white peptide components were separated:<3 k Da(PLMW),3-10 k Da(PMMW),>10 k Da(PHMW).After that,the antioxidant activities of different components were evaluated by in vitro chemical methods and an oxidative stress model of Caco-2 cells,and the highest active component was screened,and the antioxidant activity of the highest active ingredients in Caenorhabditis elegans was evaluated by using Caenorhabditis elegans model.The main research contents,results,and conclusions are as follows:(1)Egg white peptides were prepared by fermentation-assisted enzymatic hydrolysis.Single-factor tests were conducted by setting four factors: fermentation time,fermentation temperature,p H value of fermentation broth,and proportion of lactic acid bacteria.Ferric reducing antioxidant power(FRAP)assay was used as the evaluation index of the antioxidant activity of egg white peptides,and the three factors with the most significant factors on fermentation(fermentation time,fermentation temperature,and p H value of fermentation broth)were selected for further optimization of the response surface test.The optimal process conditions for fermentation-assisted preparation of egg white peptides were determined by using the three-factor three-level response surface method,of which the fermentation time was 6.25 h,the fermentation temperature was 33.27 ℃,and the p H value of the fermentation broth was 5.33.Under these conditions,the FRAP value of egg white peptide was 147.37 ± 0.807 μmo L TE·DW,which was 4.65 times that of unfermented egg white enzymatic hydrolytic peptide,indicating that the fermentation-assisted preparation of egg white peptide was effective.(2)Three kinds of ultrafiltration components,PLMW,PMMW,and PHMW,were obtained by ultrafiltration fermentation of egg white peptides with interception molecular weights of 3 k Da and 10 k Da,respectively.The antioxidant activities of different ultrafiltration components were evaluated using DPPH radical scavenging capacity,ABTS radical scavenging capacity,and iron ion reduction/antioxidant capacity.Results showed that PLMW had a high DPPH radical scavenging capacity(80.05 ±0.17%),ABTS radical scavenging capacity(82.92 ± 0.72%),and FRAP capacity(208.31 ± 1.95 μmol/TE·DW).Compared with PLMW,the antioxidant activity of PMMW was slightly weaker.The DPPH scavenging capacity,ABTS radical scavenging capacity,and FRAP capacity were 79.91 ± 0.78%,81.12 ± 0.19%,and179.49 ± 2.42 μmol/TE·DW,respectively.The antioxidant activity of PHMW ingredients was the weakest,and the scavenging capacity of DPPH,ABTS radical,and FRAP capacity were 62.70 ± 2.63%,70.30 ± 1.17%,and 91.32 ± 0.40 μmol/TE·DW,respectively.(3)By constructing the oxidative stress damage model of Caco-2 cells induced by hydrogen peroxide,the effects of different egg white peptide components on oxidative stress damage of Caco-2 cells and the contents of Superoxide Dismutase(SOD)and Reduced glutathione(GSH)in Caco-2 cells were investigated.The results showed that80 μg/m L PLMW could resist the activity damage of Caco-2 cells caused by hydrogen peroxide,and the survival rate of the cells with oxidative stress exceeded 100%.After Caco-2 cells were incubated with different components,the highest SOD and GSH contents in the cells were PLMW,which were 0.51 ± 0.04 U/mg and 34.69 ± 0.61 μg/mg prot,respectively.Followed by PMMW,with the intracellular SOD and GSH contents being 0.42 ± 0.02 U/mg and 33.09 ± 0.23 μg/mg prot,respectively.And the lowest SOD and GSH contents in the cells were PHMW,which were 0.31 ± 0.01 U/mg and 23.23 ±0.23 μg/mg prot,respectively.(4)The PLMW component with the strongest antioxidant activity in vitro was chosen to feed Caenorhabditis elegans(C.elegans),and the antioxidant activity of PLMW in vivo was evaluated by measuring the body length,lipofuscin content,reactive oxygen species(ROS)level,GSH level,superoxide anion level,and antioxidant-related enzyme activity in C.elegans.The results showed that compared with the control group,the body length of C.elegans was significantly increased by 2.59% on average after 1mg/m L PLMW feeding.Metabolites of lipofuscin content,ROS level,and superoxide anion level produced by oxidation in C.elegans showed a significant decrease trend,and they were decreased by 16.47%,23.25%,and 19.23%,respectively.At the same time,the level of GSH and the content of antioxidant-related enzymes in C.elegans increased significantly,among which the GSH level increased by 21.93%,the Peroxisome(POD)enzyme activity increased by 18.43%,and the Catalase(CAT)increased by 17.66%.1 mg/m L PLMW can significantly increase the body length of C.elegans,enhance the antioxidant capacity of the body,and have good antioxidant activity in vivo.